We modify and apply a nonlethal technique for rapidly quantifying the cold tolerance of large numbers of Drosophila and other small insects. Flies are transferred to individual vials, cooled in groups in progressive 0.5 degrees C steps, and checked for loss of righting response (chill-coma temperature [T(cc)]). Flies recover quickly when transferred to ambient temperature, and thus this technique potentially can be used in selection experiments. We applied this technique in several experiments. First, we examined the sensitivity of T(cc) to developmental temperature. Drosophila melanogaster (Congo, France), Drosophila subobscura (Spain, Denmark), and Drosophila ananassae (India) were reared from egg to adult at 15 degrees, 18 degrees, 25 degrees, or 29 degrees C, transferred to 15 degrees C for several days, and then progressively chilled: T(cc) was positively related to developmental temperature, inversely related to latitude of the population, but independent of sex. The sensitivity of T(cc) to developmental temperature (acclimation flexibility) was marked: T(cc) shifted on average 1 degrees for each 4 degrees C shift in developmental temperature. Among 15 species of the obscura group of Drosophila, T(cc) varied from -0.1 degrees to 4.5 degrees C; T(cc) was inversely related to latitude in both nonphylogenetic and phylogenetically based ANCOVA (standardized independent contrasts) and was unrelated to body size.